Regulation Of Tumor Immune Microenvironment Research Articles

Prognostic significance and response to immune checkpoint inhibitors of RIPK3, MLKL and necroptosis in non-small cell lung cancer

Lung cancer remains the leading cause of cancer death. Treatment with immune checkpoint inhibitor (ICI) alone or combination with chemotherapy served as first-line therapy in non-small cell lung cancer (NSCLC). However, only 20–50% of NSCLC patients respond to ICI. Necroptosis, an inflammatory form of cell death plays an important role in the regulation of tumor immune microenvironment which may affect prognosis and ICI response but its clinical significance in NSCLC patients has remained largely unknown. Therefore, we aimed to analyze the correlation between key necroptotic proteins and necroptosis and clinical outcomes, the status of tumor-infiltrating immune cells, and response to ICI in NSCLC patients. The expression of receptor-interacting protein kinase 3 (RIPK3), mixed lineage kinase domain-like protein (MLKL) and phosphorylated MLKL (pMLKL) were immunolocalized in 125 surgically resected NSCLC patients and 23 NSCLC patients administered with ICI therapy. CD8 + and FOXp3 + T cells and CD163 + M2 macrophages were also immunolocalized. High RIPK3 status was positively correlated with survival of the patients and RIPK3 turned out an independent favorable prognostic factor of the patients. RIPK3 was negatively correlated with CD8 + T cells, while MLKL positively correlated with CD163 + M2 macrophages, suggesting the possible involvement of RIPK3 and MLKL in formulating immunosuppressive microenvironment. In addition, high RIPK3 status tended to be associated with clinical resistance to ICI therapy (P-value = 0.057). Furthermore, NSCLC cells-expressing RIPK3 suppressed T cells response to ICI therapy in vitro. Therefore, RIPK3 and MLKL could induce an immunosuppressive microenvironment, resulting in low response to ICI therapy in NSCLC.

The clinical significance of PD-1 expression in patients with bladder cancer without lymph node metastasis: a comparative study with drained lymph nodes and tumor tissues

ABSTRACT Objective In light of the increasing importance of immunotherapy in bladder cancer treatment, this study is aim to investigate the expression and clinical significance of programmed cell surface death-1 (PD-1) in bladder cancer patients without lymph node metastasis, and to compare and analyze the difference of PD-1 in draining lymph nodes and tumor tissues. Methods The expression of PD-1 on T cells and the proportion of positive PD-1 + T cells of IFN-γ and CD105a were detected by flow cytometry, and the correlation between PD-1 expression and clinical parameters was analyzed. Results The percentage of PD-1 positive cells in drainage lymph nodes was higher than that in tumor tissues (P < 0.001). PD-1 positive cells accounted for the highest proportion in CD3 + T cells. The proportion of IFN-γ-positive PD-1 + T cells in draining lymph nodes was significantly higher than that in tumor tissues (P < 0.001), while there was no significant difference in CD105a positive PD-1 + T cells between tumor tissues and draining lymph nodes. Pathological grade, tumor size and stage were positively correlated with PD-1 expression level in the lymph nodes. Conclusion The high expression of PD-1 in patients with bladder cancer without lymph node metastasis, especially in draining lymph nodes, suggests that PD-1 may play a key role in the regulation of tumor immune microenvironment. The correlation between PD-1 and clinical parameters indicates its potential prognostic value. These findings provide important clinical implications for PD-1 targeted therapy, but further prospective studies are needed to determine the application value of PD-1 in therapeutic strategies.

Abstract B036: Micromechanical regulation of tumor immune microenvironment

Abstract Although the prognosis of primary breast cancer (BC) is considerably good, the prognosis of therapy-resistant, metastatic BC remains rather poor, thus being the major cause of BC-related deaths. Due to the undifferentiated and heterogenous nature of this disease, immunotherapies have been among the most promising therapeutic strategies for metastatic BC. However, in recent clinical trials, immunotherapies have had a poor success rate for breast cancer. Therefore, new therapeutic strategies and more profound knowledge of BC-specific immune-evasion mechanisms are urgently needed. We have developed a patient-derived explant model (PDEC), that allows us to cultivate patient-derived primary tumor samples. Moreover, the model also maintains alive the tumor resident immune cells, hence providing us with a unique platform for researching immunological drugs in vivo. In our model, tumor cells and tumor-derived immune cells are embedded in a 3D scaffold matrix with altering biological and physical properties. By altering the material in which the tumor cells are embedded we can also research the effect of microenvironmental factors such as stiffness and growth factors on the tumor cells. Microenvironmental stiffness is a well-known regulator of BC identity; in stiff 3D microenvironment breast cancer cells maintain their original identity, while in a softer microenvironment, they dedifferentiate towards more undifferentiated, therapy-resistant, and metastasis-mimicking subtype (Munne et al, 2021). In this study, we observed for the first time how the alterations in breast tumor microenvironmental stiffness also alter the phenotype in the breast tumor immune microenvironment concomitantly. In the soft microenvironment, the cytokine profile is polarized towards more tumor suppressing via upregulation of TGFB and downregulation of IL1B. Moreover, immune response and antigen presentation are heavily downregulated. Lastly, macrophages are depolarized from M1-like toward more tumor-promoting. This effect is induced and mediated by tumor cells and fresh immune cells embedded in similar 3D conditions that fail to secrete tumor suppressive cytokines or silence antitumor immunity. Manipulation of the tumor cell identity and tumor immune microenvironment via alteration of 3D matrices provides us a unique perspective for observing new methods of how metastatic breast cancer creates immune evasion. Moreover, the model allows us to develop novel treatments against metastatic, theraphy resistant breast cancer and test them easily in ex vivo conditions. Citation Format: Aino Peura, Rita Turpin, Maria Salmela, Ruixian Liu, Piia Mikkonen, Juha Klefström, Pauliina M Munne. Micromechanical regulation of tumor immune microenvironment [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr B036.

A novel prognostic signature and immune microenvironment characteristics associated with disulfidptosis in papillary thyroid carcinoma based on single-cell RNA sequencing.

Background: Disulfidptosis is a newly discovered form of regulated cell death. The research on disulfidptosis and tumor progression remains unclear. Our research aims to explore the relationship between disulfidptosis-related genes (DRGs) and the clinical outcomes of papillary thyroid carcinoma (PTC), and its interaction on the tumor microenvironment. Methods: The single-cell RNA seq data of PTC was collected from GEO dataset GSE191288. We illustrated the expression patterns of disulfidptosis-related genes in different cellular components in thyroid cancer. LASSO analyses were performed to construct a disulfidptosis associated risk model in TCGA-THCA database. GO and KEGG analyses were used for functional analyses. CIBERSORT and ESTIMATE algorithm helped with the immune infiltration estimation. qRT‒PCR and flow cytometry was performed to validate the hub gene expression and immune infiltration in clinical samples. Results: We clustered PTC scRNA seq data into 8 annotated cell types. With further DRGs based scoring analyses, we found endothelial cells exhibited the most relationship with disulfidptosis. A 4-gene risk model was established based on the expression pattern of DRGs related endothelial cell subset. The risk model showed good independent prognostic value in both training and validation dataset. Functional enrichment and genomic feature analysis exhibited the significant correlation between tumor immune infiltration and the signature. The results of flow cytometry and immune infiltration estimation showed the higher risk scores was related to immuno-suppressive tumor microenvironment in PTC. Conclusion: Our study exhibited the role of disulfidptosis based signature in the regulation of tumor immune microenvironment and the survival of PTC patients. A 4-gene prognostic signature (including SNAI1, STC1, PKHD1L1 and ANKRD37) was built on the basis of disulfidptosis related endothelial cells. The significance of clinical outcome and immune infiltration pattern was validated robustly.

(E)-SIS3 suppressed osteosarcoma progression via promoting cell apoptosis, arresting cell cycle, and regulating the tumor immune microenvironment.

Osteosarcoma is a prevalent malignant bone tumor with a poor prognosis. Mothers against decapentaplegic homolog 3 (Smad3) present as a therapeutic target in antitumor treatment, whereas its functions in the osteosarcoma have not been well explored. In the current study, we aimed to investigate the effects of Smad3 in the progression of osteosarcoma. The tumor immune single-cell hub 2 website was used for graph-based visualization of Smad3 status in osteosarcoma single-cell database. Western Blot was applied to detect the expression of Smad3 protein in cell lines. Colony formation and cell counting kit-8 assays were used to evaluate cell proliferation. Transwell and wound healing assays were used to detect the migration and invasion abilities of cells. Cell apoptosis rates and cell cycle changes were explored by using flow cytometry analysis. The xenograft tumor growth model was applied to explore the effect in tumor growth after Smad3 blockage in vivo. Moreover, to confirm the potential mechanism of Smad3's effects on osteosarcoma, bioinformatics analysis was performed in TARGET-Osteosarcoma and GSE19276 databases. Our study found that the Smad3 protein is overexpressed in 143B and U2OS cells, suppressing the expression of Smad3 protein in osteosarcoma cells by Smad3 target inhibitor (E)-SIS3 or lentivirus can inhibit the proliferation, migration, invasion, promote cell apoptosis, arrest cell G1 cycle in osteosarcoma cells in vitro, and suppress tumor growth in vivo. Furthermore, the bioinformatics analysis demonstrated that high expression of Smad3 is closely associated with low immune status in TARGET-Osteosarcoma and GSE19276 databases. Our study suggested that Smad3 could contribute positively to osteosarcoma progression via the regulation of tumor immune microenvironment, and Smad3 may represent as an valuable potential therapeutic target in osteosarcoma therapy.

Artificial nanoplatelet regulation of tumor immune microenvironment to inhibit post-surgical tumor recurrence and lung metastasis

Tumor recurrence and metastasis are important causes of death in breast cancer patients after surgery. However, the nanomedicines are often difficult to accumulate and retain in residual microtumors because the surgery disrupts the original network of blood vessels in the solid tumor, which is a crucial factor leading to tumor recurrence and metastasis after surgery. Herein, an engineered artificial nanoplatelet (MOAP) was established by loading antigen protein ovalbumin (OVA) and PD-L1 inhibitor (aptamer-PD-L1) on MnOx nanoflowers and further coating with platelet membrane. MOAP can mimic platelets to participate in the coagulation process, target thrombus at the residual tumor site, and release OVA and Apt-PD-L1 under 808 nm laser irradiation to block immune checkpoint inhibition and prevent immune escape. Moreover, MnOx alleviated hypoxia, promoted the polarization of tumor-associated macrophages from M2 to M1 phenotype, and significantly reversed the tumor immunosuppressive microenvironment, thus enhancing the infiltration of cytotoxic T lymphocytes at the tumor site. In vivo experiments revealed that pre-surgery or post-surgery treatment of artificial platelets had a good anti-tumor effect, and inhibited tumor recurrence and lung metastasis after surgery, which demonstrated that MOAP is a suitable system for the treatment of post-surgery tumors.

Cancer Cell Membrane-Coated Gambogic Acid Nanoparticles for Effective Anticancer Vaccination by Activating Dendritic Cells.

Recent studies have shown that traditional Chinese medicine (TCM), such as gambogic acid (GA), is involved in the regulation of tumor immune microenvironment and can be combined with other anti-tumor treatment strategies. Here, we used GA as an adjuvant to construct a nano-vaccine to improve the anti-tumor immune response of colorectal cancer (CRC). We used a previously reported two-step emulsification method to obtain poly (lactic-co-glycolic acid) /GA nanoparticles (PLGA/GA NPs), and then CT26 colon cancer cell membrane (CCM) was used to obtain CCM-PLGA/GA NPs. This novel nano-vaccine, CCM-PLGA/GA NPs, was co-synthesized with GA as an adjuvant and neoantigen provided by CT26 CCM. We further confirmed the stability, tumor targeting, and cytotoxicity of CCM-PLGA/GA NPs.The regulatory effect on the tumor immune microenvironment, the anti-tumor efficacy, and the combined anti-tumor efficacy with anti-PD-1 monoclonal Antibodies (mAbs) of this novel nano-vaccine was also detected in vivo. We successfully constructed the CCM-PLGA/GA NPs. In vitro and in vivo tests showed low biological toxicity, as well as the high tumor-targeting ability of the CCM-PLGA/GA NPs. Besides, we revealed a remarkable effect of CCM-PLGA/GA NPs to activate the maturation of dendritic cells (DCs) and the formation of a positive anti-tumor immune microenvironment. This novel nano-vaccine constructed with GA as the adjuvant and CCM providing the tumor antigen can not only directly kill tumors by enhancing the ability of GA to target tumors, but also indirectly kill tumors by regulating tumor immune microenvironment, providing a new strategy for immunotherapy of CRC.

Aspartate beta-hydroxylase domain containing 1 as a prognostic marker associated with immune infiltration in skin cutaneous melanoma

BackgroundSkin cutaneous melanoma (SKCM) is an extremely malignant tumor and accounts for the majority of skin cancer deaths. Aspartate beta-hydroxylase domain containing 1 (ASPHD1) may participate in cancer progression through controlling α-ketoglutarate-dependent dioxygenases. However, its role in skin cutaneous melanoma (SKCM) has not been well studied.MethodsThe gene expression data of ASPDH1 and differentially expressed genes (DEGs) from TCGA and GTEx were evaluated, and verified via the GEO database. Then, we performed GO/KEGG, GSEA, PPI network analysis to analyze the functional implications of the DEGs related to ASPHD1. Then, the association between the ASPHD1 expression and clinical parameters was investigated by Cox regression analysis. Subsequently, the survival time of SKCM patients was evaluated by plotting Kaplan-Meier curves. Moreover, we investigated the correlation between the ASPHD1 expression and lymphocytic infiltration by using the data from TISIDB and TIMER 2.0. Next, we explored the association between ASPHD1 expression and drug sensitivity. Finally, we validate the expression differences by analyzing the results of qPCR, Western blot from human normal epidermal melanocytes and melanoma cells, and immunohistochemistry (IHC) from non-tumor skin as well as melanoma tissues.ResultsThe ASPHD1 expression level was significantly upregulated in several cancers, including SKCM especially SKCM-metastasis tissues, and patients with an increased ASPHD1 expression had longer overall survival time than low expression ones. The functional enrichment analysis of ASPHD1-related DEGs showed an association with cell development regulation and tumorigenic pathways. Furthermore, the increased ASPHD1 expression level was associated with the level of immunostimulors, immunoinhibitors, chemokines, and TILs, such as CD4+, CD8+ T cell, mast cell, Th2 cell, and dendritic cell. More interesting, we found that ASPHD1 expression was tightly associated with CTLA4 and CD276 which are immune checkpoint markers. Moreover, the upregulated expression of ASPHD1 exhibited higher IC50 values for 24 chemotherapy drugs, including doxorubicin, and masitinib. Finally, the differential expression of ASPHD1 in SKCM was validated by the results of qPCR, Western blot, and IHC.ConclusionThe expression of ASPHD1 in SKCM patients is closely related to patient survival. ASPHD1 may participate in the regulation of tumor immune microenvironment. Additionally, it may serve as a prognostic biomarker for SKCM and future in-depth studies are necessary to explore its value.

Anti-tumor effects of miR-34a by regulating immune cells in the tumor microenvironment.

Malignant tumors pose a serious threat to human health. The development of malignant tumors is characterized by uncontrolled cell division and immune evasion. The micro-ribonucleic acid-34a (miR-34a) is a small noncoding single-stranded ribonucleic acid that is ubiquitously present in normal human tissues. However, it has been confirmed to be dysregulated in a variety of tumor cells. Numerous research have revealed the importance of miR-34a in the treatment of various malignancies. MiR-34a deletion can hasten the growth of tumors whereas miR-34a overexpression suppresses the proliferation, invasion, and migration of cancer cells. Moreover, more recent studies have highlighted its role in immunity and investigated its applicability to particular tumors. Through various immune cells, factors, and other mechanisms, miR-34a can inhibit tumor carcinogenesis. In view of the important role of miR-34a in tumors, this research reviewed the aspects of miR-34a regulation of tumor immune microenvironment to exert anti-tumor effects in order to clarify the potential immunotherapy value of miR-34a in tumors.

Combination of a cationic complexes loaded with mRNA and α-Galactose ceramide enhances antitumor immunity and affects the tumor immune microenvironment

mRNA vaccination is considered to be a promising strategy for tumor immunotherapy. Among, adequate antigen expression and regulation of tumor immune microenvironment are still the key to achieving therapeutic immounotherapy. In oreder to protect mRNA delivered to cells and reverse damaged dendritic cells(DCs), a novel vaccine delivery system composed of an α-Galactose ceramide/cationic liposome complex(α-GC-Lip) was constructed. The α–GC-liposome/protamine/mRNA vaccine complexes(α-GC-LPR) enabled the mRNA to be successfully translated into protein in the cytoplasm of antigen-presenting cells. Further, α-GC-LPR could stimulate dendritic cell maturation via significantly increasing the expression of bone marrow-derived cells(BMDCs) surface molecules and secretion of cytokines to improve the efficacy of immunotherapy. In vivo study, the α-GC-LPR was combined with programmed cell death protein 1(PD-1) inhibitor could activate natural killer cell(NK), T cells as well as significantly reduce the immunosuppression of immune cells, which induced strong antigen-specific immunity in breast cancer model. Our study indicated that the α-GC-LPR combined with immune checkpoint inhibitors as a potential design strategy to effectively enhance the antitumor immune response.

Silver nanoclusters show advantages in macrophage tracing in vivo and modulation of anti-tumor immuno-microenvironment.

Macrophage-based nanomedicine represents an emerging powerful strategy for cancer therapy. Unfortunately, some obstacles and challenges limit the translational applications of macrophage-mediated nanodrug delivery system. For instance, tracking and effective cell delivery for targeted tumor sites remain to be overcome, and controlling the states of macrophages is still rather difficult due to their plastic nature in response to external stimuli. To address these critical issues, here, we reported a novel type of silver nanoclusters (AgNCs) with excellent fluorescent intensity, especially long-lasting cell labeling stability after endocytosis by macrophages, indicating promising applications in tracking macrophage-based nanomedicine delivery. Our mechanistic investigations uncovered that these merits originate from the escape of AgNCs from lysosomal degradation within macrophages. In addition, the AgNCs would prime the M1-like polarization of macrophages (at least in part) through the toll-like receptor 4 signaling pathway. The engineered macrophages laden with AgNCs could be employed for lung metastasis breast cancer treatment, showing the effective targeting propensity to metastatic tumors, remarkable regulation of tumor immune microenvironment and inhibition of tumor growth. Collectively, AgNC-trained macrophages appear to be a promising strategy for tumor immune-microenvironment regulation, which might be generalized to a wider spectrum of cancer therapeutics.

Regulation of Tumor Immune Microenvironment by Sphingolipids and Lysophosphatidic Acid

The tumor microenvironment (TME) consists of cancer cells that interact with stromal components such as the extracellular matrix, blood, and lymphatic networks, fibroblasts, adipocytes, and the cells of the immune system. Further, the tumor immune microenvironment, majorly represented by the tumor-infiltrating immune cells (TIIC), plays an important role in cancer therapeutics and patient prognosis. In fact, a high density of TIICs within the tumor microenvironment is known to be associated with better outcomes in several types of cancers. Towards this, two bioactive lipid molecules, lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P), regulate the homing of immune cells to the TME. In the present review, we will uncover the role of LPA and S1P signaling in the tumor immune environment, highlighting the latest progress in this field.

EEF2K promotes PD-L1 stabilization through inactivating GSK3β in melanoma

BackgroundImmune checkpoint blockade (ICB) targeting programmed death ligand-1 (PD-L1)/programmed cell death protein-1 (PD-1) pathway has become an attractive strategy for cancer treatment; however, unsatisfactory efficacy has limited its clinical benefits....

A Pyroptosis-Based Prognostic Model for Immune Microenvironment Estimation of Hepatocellular Carcinoma.

Background Hepatocellular carcinoma (HCC), an aggressive malignant tumor, has a high incidence and unfavorable prognosis. Recently, the synergistic effect of pyroptosis in antitumor therapy and regulation of tumor immune microenvironment has made it possible to become a novel therapeutic method, but its potential mechanism still needs further exploration. Methods Differentially expressed genes with prognostic value in Liver Hepatocellular Carcinoma Project of The Cancer Genome Atlas (TCGA-LIHC) cohort were screened and incorporated into the risk signature by Cox proportional hazards regression model and least absolute shrinkage and selection operator. Kaplan-Meier (KM) curves and receiver operating characteristic (ROC) curves were applied to conduct survival comparisons and estimate prediction ability. The dataset of Liver Cancer-RIKEN, Japan Project from International Cancer Genome Consortium (ICGC-LIRI-JP) cohort was used to verify the reliability of the signature. Correlation analysis between clinicopathological characteristics, immune infiltration, drug sensitivities, and risk scores was conducted. Functional annotation analyses were performed for the genes differentially expressed between high-risk and low-risk groups. Results A risk signature consisting of 6 pyroptosis-related genes in HCC was developed and validated. KM curves and ROC curves revealed its considerable predictive accuracy. Higher risk scores meant more advanced grade, higher alpha-fetoprotein level, and stronger invasive ability. Overexpressed genes in high-risk population were more enriched in the immune-associated pathways, and these patients might be more sensitive to immune checkpoint inhibitors instead of Sorafenib. Intriguingly, 6 identified genes were promising to be prognostic biomarkers and therapeutic targets of HCC. Conclusions The signature may have crucial clinical significance in predicting survival prognosis, immune infiltration, and drug efficacy based on pyroptosis-related genes.

Immune Checkpoint Inhibitors in Hepatocellular Carcinoma: Current Progresses and Challenges.

Hepatocellular carcinoma (HCC) is one of the most common malignant tumor in the world and its incidence is increasing in many countries. In recent years, with the deepening understanding of the immune and pathological mechanisms of HCC, immunotherapy based on the regulation of tumor immune microenvironment has become a new treatment choice for patients with HCC. Immune checkpoint inhibitors (ICIs) targeting programmed death protein-1, programmed death protein-ligand-1, or cytotoxic T-lymphocyte-associated antigen 4 are the most widely used. Instead of general immune-enhancing therapies, ICIs can reactivate anti-tumor immune responses by disrupting co-inhibitory T cell signaling. In this review, the research progress and existing problems of ICIs in the treatment of HCC in recent years are reviewed.

KAT2B is an immune infiltration-associated biomarker predicting prognosis and response to immunotherapy in non-small cell lung cancer.

Over the past few years, dramatic breakthroughs in the field of tumor immunotherapy with immune checkpoint inhibitors (ICIs) have made a therapeutic revolution for non-small cell lung cancer (NSCLC). While only some patients present a favorable response to this treatment. It is urgent to explore the potential molecular mechanisms underlying the regulation of tumor immune microenvironment in the process of immunotherapy. Lysine acetyltransferase 2B (KAT2B) plays a crucial role in the regulation of gene expression at the post-transcriptional level by acetylation, and is associated with many types of cancer. RNA-sequencing data,genetic mutation data,and corresponding clinical information were extracted from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases, then subjected to immune characteristics, gene expression, survival, genetic alteration, enrichment analyses. KAT2B expression correlated positively with infiltrating levels of multiple immune cells and mRNA expression levels of immune checkpoint genes in NSCLC. Furthermore, KAT2B expressionwas downregulated in tumor tissues, and low KAT2B expression was associated with unsatisfactory efficacy of immune checkpoint blockade (ICB) and poor prognosis of patients with lung adenocarcinoma. Moreover, there were higher somatic genes mutation frequency in patients with low expression of KAT2B. Finally, functional enrichment analysis suggested that KAT2B was mainly linked to the regulation of immune cells and interferon - gamma (IFN-γ) mediated signaling pathways, response to IFN-γ, antigen processing and presentation. This is the first comprehensive study to disclose that KAT2B is correlated with immune infiltrates and may serve as a novel biomarker predicting prognosis and response to immunotherapy in NSCLC.

Comprehensive Analysis of the Clinical and Biological Significances of Endoplasmic Reticulum Stress in Diffuse Gliomas.

BackgroundAs a critical organelle for protein and lipid synthesis, the dysfunction of endoplasmic reticulum has a significant impact on multiple biological processes of cells. Thus, in this study, we constructed an ER stress-related risk signature to investigate the functional roles of ER stress in gliomas.MethodsA total of 626 samples from TCGA RNA-seq dataset (training cohort) and 310 samples from CGGA RNA-seq dataset (validation cohort) were enrolled in this study. Clinical information and genomic profiles were also obtained. The ER stress signature was developed by the LASSO regression model. The prognostic value of the risk signature was evaluated by Cox regression, Kaplan-Meier and ROC Curve analyses. Bioinformatics analysis and experiment in vitro were performed to explore the biological implication of this signature.ResultsWe found that the ER stress-related signature was tightly associated with major clinicopathological features and genomic alterations of gliomas. Kaplan-Meier curve and Cox regression analysis indicated that ER stress activation was an independent prognostic factor for patients with glioma. Besides, we also constructed an individualized prognosis prediction model through Nomogram and ROC Curve analysis. Bioinformatics analysis suggested that ER stress activation also promoted the malignant progression of glioma and participated in the regulation of tumor immune microenvironment, especially the infiltration of macrophages in M2 phase. These results were further validated in IHC analysis and cell biology experiments.ConclusionThe ER stress activation had a high prognostic value and could serve as a promising target for developing individualized treatment of glioma.

Abstract 439: BCL-2 inhibitor APG-2575 promotes anti-tumor efficacy of immune checkpoint blockade through converting tumor-associated macrophages into M1 phenotype in non-small cell lung cancer

Abstract BCL-2 inhibitor APG-2575 promotes anti-tumor efficacy of immune checkpoint blockade through converting tumor-associated macrophages into M1 phenotype in non-small cell lung cancer Background: Immune checkpoint inhibitors (ICIs) has received approval due to its encouraging clinical efficacy with improving the outcome of cancer patients. However, current immunotherapeutic regimens benefit merely a minority of populations. Thus, combining ICIs with other potential modalities is of great significance to enhance the response of single drug alone. Modifying tumor-associated macrophages (TAMs) strategies have attracted attention for ameliorating tumor immunosuppression and improving anti-tumor immunity. Here we exhibit APG2575, a proven BCL-2 inhibitor, can act as an antitumor immune modulator that resets TAMs from M2 to pro-tumoral M1 phenotype and further augment immunotherapeutic efficacy. Methods: By applying non-small cell lung cancer cell lines and murine syngeneic tumor xenograft, we investigated the effects of combined APG-2575 and PD-1 antibody on tumor growth. Furthermore, we explored the role of APG-2575 in regulating TAMs polarization through utilizing human and murine macrophages, involving immortalized and primary models. Besides, IL-10 secretion in cell culture supernatants was detected via ELISA assay. In addition, we also assessed the tumor immune microenvironment regulation of APG-2575 using flow cytometry. Results: In vitro, APG-2575, a BCL-2 inhibitor, augmented the tumor cell lysis induced by T cells when combined with ICIs. Moreover, mice treated with APG-2575 and ICIs showed remarkable tumor regression and prolonging survival which were associated with the increase of granzyme-B, IFN-γ and TNF-α release of CD8+ cytotoxic lymphocytes. Importantly, we found that APG-2575-mediated antitumor T cell immunity is dependent on macrophages. Furthermore, we discovered APG-2575 could induce M2 macrophage switching to M1 phenotype and suppress IL-10 secretion in vitro. Meanwhile, reduced M2 macrophages as well as increased M1 polarization were also observed in the tumor-infiltrating immune cells from mice treated with APG-2575. Additionally, APG-2575-mediated macrophage transition could improve tumor immunosuppression, with decreasing number of myeloid-derived suppressor cells (MDSCs) and regulatory T (Treg) cells in tumor microenvironment. Conclusions: Our data demonstrate that APG-2575 can synergizes with ICIs in vitro and in vivo, which was involved in switching TAMs from M2 to M1 polarization and further improving tumor immunosuppression, finally enhancing T cell anti-tumor activity. These results warranted future clinical evaluation of combination therapy of APG-2575 and ICIs. Key words: BCL-2, APG-2575, ICIs, macrophages, non-small cell lung cancer Citation Format: Fan Luo, Fei Teng Lu, Miao Zhen Qiu, Wen Tao Pan, Lin Zhang, Hong Yun Zhao, Li Zhang, Da Jun yang. BCL-2 inhibitor APG-2575 promotes anti-tumor efficacy of immune checkpoint blockade through converting tumor-associated macrophages into M1 phenotype in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 439.

NEFM DNA methylation correlates with immune infiltration and survival in breast cancer

BackgroundThis study aims to determine whether NEFM (neurofilament medium) DNA methylation correlates with immune infiltration and prognosis in breast cancer (BRCA) and to explore NEFM-connected immune gene signature.MethodsNEFM transcriptional expression was analyzed in BRCA and normal breast tissues using Oncomine and Tumor Immune Estimation Resource (TIMER) databases. The relationship between NEFM DNA methylation and NEFM transcriptional expression was investigated in TCGA. Potential influence of NEFM DNA methylation/expression on clinical outcome was evaluated using TCGA BRCA, The Human Protein Atlas and Kaplan–Meier plotter databases. Association of NEFM transcriptional expression/DNA methylation with cancer immune infiltration was investigated using TIMER and TISIDB databases.ResultsHigh expression of NEFM correlated with better overall survival (OS) and recurrence-free survival (RFS) in TCGA BRCA and Kaplan–Meier plotter, whereas NEFM DNA methylation with worse OS in TCGA BRCA. NEFM transcriptional expression negatively correlated with DNA methylation. NEFM DNA methylation significantly negatively correlated with infiltrating levels of B, CD8+ T/CD4+ T cells, macrophages, neutrophils and dendritic cells in TIMER and TISIDB. NEFM expression positively correlated with macrophage infiltration in TIMER and TISIDB. After adjusted with tumor purity, NEFM expression weekly negatively correlated with infiltration level of B cells, whereas positively correlated with CD8+ T cell infiltration in TIMER gene modules. NEFM expression/DNA methylation correlated with diverse immune markers in TCGA and TISIDB.ConclusionsNEFM low-expression/DNA methylation correlates with poor prognosis. NEFM expression positively correlates with macrophage infiltration. NEFM DNA methylation strongly negatively correlates with immune infiltration in BRCA. Our study highlights novel potential functions of NEFM expression/DNA methylation in regulation of tumor immune microenvironment.

CD3D has the Potential to be a Prognostic Factor for Endometrial Carcinoma and an Indicator of Tumor Immune Microenvironment Regulation: a Study based on TCGA Data Mining

Accumulating evidence shows that tumor immune microenvironment infiltration plays a crucial role in the occurrence and development of endometrial carcinoma (EC). At present, studying the regulation mechanism of the immune microenvironment in EC remains a challenge. We calculated the amount of immune and stromal components and the proportion of tumor-infiltrating immune lymphocytes via ESTIMATE, single-sample gene enrichment analysis (ssGSEA), and CIBERSORT computational methods. Differentially expressed genes were analyzed by protein–protein interaction network construction and univariate survival analysis. CD3D expression was negatively correlated with clinicopathological features, such as age, histological type, clinical grade, and pathological stage, but positively correlated with the survival of patients with EC. Logistic regression analysis suggested that CD3D expression could be used as a biomarker to predict EC occurrence. Gene set enrichment analysis revealed that CD3D was upregulated in the immune effect pathway. Interestingly, CD3D was downregulated in the EC pathway. EC was divided into three immune cell subsets by ssGSEA. Correlation analysis between ssGSEA and CIBERSORT showed that the infiltration level of CD8 + T cells and plasma cells in tumor-infiltrating lymphocytes was lower than that in the high-immune-score group (Immunity_H). Moreover, CD3D expression in Immunity_H was low, and the prognosis was even worse. This study demonstrated that low CD3D expression is an index of poor prognosis in patients with EC. Levels of CD3D in tumor microenvironment (TME) might be a useful indicator for predicting the prognosis of patients with EC patients.